Project Summary As the U.S. and global populations continue to age, a concomitant increase disease is expected. The increase will be in multiple diverse diseases, including post-surgical, post-traumatic, oncologic, and degenerative diseases. Ultimately, the tissue response to disease and injury is fibrosis and scarring, which degrades tissue function. The cost of this biomedical burden resulting from lost tissue function is enormous. Alternative approaches for treatment of tissue scar and fibrosis must be developed, as no effective treatment exists. The skin is an ideal system for understanding tissue regeneration as skin responds to injury with perfect regeneration during fetal development. Understanding the mechanisms of skin regeneration in the fetus will lead to development of regenerative treatment capabilities in the adult. Recent studies have focused on a variety of wound healing mechanistic differences between scarless fetal wounds and scarring adult wounds. These investigations range from growth factor expression, extracellular matrix deposition, and inflammation differences. However the responsible mechanism(s) has not been identified. Our preliminary data has identified a unique dermal fibroblast lineage that is the primary contributor to connective tissue secretion and fibrotic scar formation during cutaneous wound repair. We believe that this lineage is responsible for skin scar formation after injury. By determination of fibroblast lineages based on HOX gene expression during embryonic development, we found that the Engrailed-1 (En1) lineage is the primary contributor to connective tissue secretion and organization during embryonic development and cutaneous wound repair. Our central hypothesis is that different lineages of dermal fibroblasts are responsible for scarless and scarring repair. In this proposal, we will demonstrate that the En1 lineage of dermal fibroblast cells have a more fibrotic response during repair, compared to non-lineage fibroblasts; and En1 lineage fibroblasts are responsible for the transition from scarless healing to scarring repair in skin, due to a cell-intrinsic process. We will demonstrate that targeted blocking of En1 lineage dermal fibroblast function during repair reduces scarring and induces regenerative healing. We expect that therapeutic strategies will be developed to target these cells for specific tissue regeneration after injury and disease rather than fibrotic tissue formation. We anticipate our findings to significantly move the field of regenerative medicine forward. Furthermore, our findings will support the notion that the term `fibroblast' represents a heterogeneous population of cells composed of multiple lineages, each with distinct embryonic origins, migratory routes, and functional properties. These findings of fibroblast lineage specificity will have a lasting and powerful impact as the mechanistic basis for investigation into fibroblast lineage-specific response to injury is provided.